ASCP Tissue Fixation and Decalcification 2 — Questions and Answers
Question 1: Which fixative type is most likely to mask antigenic epitopes, necessitating antigen retrieval before immunohistochemistry?
- 10% neutral buffered formalin (Correct answer)
- Absolute ethanol
- Acetone
- Methanol
Correct answer: 10% neutral buffered formalin
Formalin cross-links proteins and masks antigenic epitopes; heat-induced or proteolytic antigen retrieval must be performed to unmask these epitopes before IHC staining.
Question 2: Zenker's fixative contains which hazardous heavy metal component?
- Lead acetate
- Mercuric chloride (Correct answer)
- Arsenic trioxide
- Cadmium sulfate
Correct answer: Mercuric chloride
Zenker's fixative contains mercuric chloride, which is highly toxic and requires special handling and disposal procedures, contributing to its decline in modern laboratories.
Question 3: Which fixative is preferred when preservation of glycogen in tissue is the primary concern?
- 10% neutral buffered formalin
- Carnoy's fixative
- Absolute alcohol (ethanol) (Correct answer)
- Bouin's solution
Correct answer: Absolute alcohol (ethanol)
Absolute alcohol preserves glycogen by precipitating surrounding proteins through dehydration without dissolving the water-soluble glycogen molecules.
Question 4: What is the ideal pH range for neutral buffered formalin to prevent artifact formation?
- 5.0–6.0
- 6.0–6.8
- 6.8–7.4 (Correct answer)
- 7.4–8.0
Correct answer: 6.8–7.4
NBF should be maintained between pH 6.8 and 7.4 to prevent the formation of formalin pigment (acid formaldehyde hematin) and optimize tissue preservation.
Question 5: What is formalin pigment and how is it prevented?
- A brown artifact from acidic formalin reacting with hemoglobin; prevented by using buffered formalin (Correct answer)
- A yellow discoloration from picric acid; prevented by washing in tap water
- A blue artifact from alkaline formalin; prevented by using unbuffered formalin
- A black deposit from osmium tetroxide; prevented by shortening fixation time
Correct answer: A brown artifact from acidic formalin reacting with hemoglobin; prevented by using buffered formalin
Formalin pigment (acid formaldehyde hematin) is a brown refractile artifact that forms when acidic, unbuffered formalin reacts with hemoglobin in bloody tissue; buffering the formalin to neutral pH prevents its formation.
Question 6: What is the recommended tissue-to-fixative volume ratio to ensure adequate fixation?
- 1:1
- 1:5
- 1:10 (Correct answer)
- 1:20
Correct answer: 1:10
A 1:10 tissue-to-fixative ratio ensures sufficient fixative volume to penetrate the specimen completely and maintain fixative concentration throughout the process.
Question 7: Ethanol (absolute alcohol) is classified as which type of fixative based on its mechanism of action?
- Additive cross-linking fixative
- Non-additive coagulant fixative (Correct answer)
- Oxidizing fixative
- Metallic fixative
Correct answer: Non-additive coagulant fixative
Ethanol is a non-additive coagulant fixative that fixes tissue by precipitating and denaturing proteins through dehydration, without forming covalent cross-links.
ASCP Tissue Fixation and Decalcification 2
Which fixative type is most likely to mask antigenic epitopes, necessitating antigen retrieval before immunohistochemistry?